The “thousand dollar genome” for use in personalized medicine is a much-sought goal in DNA sequencing. Until recently the cost of sequencing the genome of an individual human was estimated to be at least $10 million. Current methods show promise of reducing the cost of sequencing a human genome to $100,000 or perhaps even $20,000. See, for example, “Genome sequencing in microfabricated high-density picolitre reactors,” by Marcel Margulies et al., Nature, Jul. 30, 2005, (advance published at nature.com/nature/journal and more specifically on the internet at http://www.nature.com/nature/journal/vaop/ncurrent/pdf/nature03959.pdf), which is incorporated herein in its entirety by reference. However, present methods show no promise of reaching the price point of $1,000, which would make sequencing of an individual's DNA feasible as a routine diagnostic procedure.
The use of synthetically fabricated nanoscale pores, called synthetic nanopores or sometimes simply nanopores, shows promise in performing rapid DNA sequencing. Such pores can in principle be used to position a sensing system, such as electrodes, in close proximity to a DNA strand passing through a nanopore. The nanopore in principle performs a sizing function, restricting molecular passage to one strand of DNA at a time, either single-stranded DNA (ssDNA) or double-stranded DNA (dsDNA). Passage of a molecule through the nanopore is typically performed by use of an applied voltage between two ionic conducting liquid volumes disposed on either side of the nanopore. The applied voltage creates an electric field through the nanopore, and the electric field acts to pull charged long-chain polymer molecules, such as DNA, through the nanopore in a chain-wise fashion, one monomer unit after another. Other molecules, such as RNA or polypeptides, can in principle also be sized by a nanopore and characterized as they pass through the nanopore.
Naturally-occurring biological nanopores (e.g., an alpha-hemolysin (α-hemolysin) molecule) have been placed in lipid bilayer membranes and employed to pass single stranded DNA. However, it is difficult to place any sensing system in association with such pores, and early plans to use such pores for DNA sequencing using ionic current measurement, as typified by Baldarelli et al. (U.S. Pat. No. 6,015,714), have proven to be ineffective because fundamental thermal noise considerations in the ionic current going through the nanopore dictate that the current change due to a single nucleotide unit is too small to detect. Still, the α-hemolysin molecule does have the advantageous characteristics of atomic precision and reproducibility, that is, each α-hemolysin molecule is the same in size, shape, and nanopore diameter. If it were larger diameter and able to be instrumented with a sensing system having better signal-to-noise characteristics, it would be more useful.
More recently, modifications of semiconductor fabrication technology have been used to fabricate synthetic nanopores with larger diameters than a-hemolysin nanopores. Synthetic nanopores fabricated to date have been formed, typically in thin membranes of silicon nitride, by a combination of focused ion beam drilling and argon ion beam sculpting as per Golovchenko, et al., in U.S. Patent Application Publication No. US 2005/0126905, but these processes are slow and inexact, producing nanopores of variable size and shape. Sorting of individual chips containing properly sized pores is possible, but at a cost that is likely too high for purposes of achieving the thousand-dollar genome. Monitoring of ionic current through such pores produces a rough length estimate for a DNA strand passing through the pore, but this sensing modality shows little hope of sequencing the molecule.
Ultra-high-throughput (UHT) DNA sequencing systems using nanopores will require the use of many synthetic nanopores in parallel, placed on one substrate for low cost, each fabricated to a desired size and shape with good uniformity. In addition, each pore must be instrumented with a sensing system having better signal-to-noise characteristics than ionic current sensing systems.
Providing electrodes to instrument the nanopore, and measure a tunneling current through DNA bases transiting the pore, has been described as a method to rapidly sequence DNA by Flory in U.S. Patent Application Publication No. US2004/0144658A1. This method offers higher current levels and better signal-to-noise ratios than the use of ionic currents through the pore, and might be incorporated into systems for UHT DNA sequencing.
One method of fabricating nanopores instrumented with resonant tunneling electrodes has been described and this method uses ion beam sculpting to fabricate the nanopore, much like that described in the aforementioned U.S. Patent Application US 2005/0126905, and as reasoned above, provides pores of variable diameter that may be too expensive for purposes of achieving the thousand-dollar genome.
In a different area of technology, fabrication of nanoscale transistors employing nanotubes or nanowires runs into difficulties. One such difficulty is in placing the source and drain electrical contacts of a nanoscale transistor adjacent, but not touching, the gate contact of the transistor. A second difficulty is in placing a gate contact so that it encircles a nanotube or a nanowire in a fashion providing low electrical capacitance.
The present disclosure addresses some of the limitations described above.